Methods to restore glycemic control

ABSTRACT

Provided herein are methods and compositions to achieve a sustained delay in the progression of, or an amelioration of diabetes in a subject, or a delay in diabetes onset in a subject at risk for diabetes, comprising an abbreviated course of administration of a pharmaceutical composition comprising an exendin or an exendin agonist analog in an amount effective to induce β cell regeneration.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.13/619,495, filed Sep. 14, 2012, which is a continuation of U.S.application Ser. No. 12/298,933, filed Jan. 7, 2009, now U.S. Pat. No.8,299,024, which is a U.S. National Stage of PCT/US07/11641, filed May14, 2007, which claims the benefit of U.S. Prov. Appl. No. 60/799,913,filed May 12, 2006; the contents of each of which are herebyincorporated by reference herein in the entirety and for all purposes.

FIELD OF THE INVENTION

The present invention relates to the fields of medicine and health. Moreparticularly, the present invention relates to methods and compositionsrestore glycemic control in subjects, for example, subjects withdiabetes.

BACKGROUND OF THE INVENTION

Diabetes mellitus is characterized by insufficiency of the pancreatic βcells to maintain normal levels of blood glucose or normoglycemia. Undernormal conditions, pancreatic β cells secrete insulin (as well as otherhormones) in response to an increased blood glucose level, e.g., after ameal. Insulin acts to lower blood glucose levels through actions thatinclude stimulation of fat synthesis, promotion of triglyceride storagein fat cells, and promotion of protein synthesis in the liver andmuscle. In diabetes, the inability to maintain normoglycemia resultsfrom a failure of the pancreatic β cells to produce insulin, thedevelopment of insulin resistance in tissues that typically participatein blood glucose regulation, or some combination of these. Thesubsequent hyperglycemia contributes significantly to an increased riskof cardiovascular disease, neuropathy, nephropathy, retinopathy,hypertension, dyslipidemia, as well as increased morbidity andmortality.

In most patients with type I diabetes, pancreatic β cells are destroyedby an autoimmune response that can result in an absolute deficiency ininsulin production. While some patients lack evidence of an autoimmuneresponse against pancreatic β cells (known as idiopathic type 1diabetics), the absence of pancreatic β cell still manifests itself as adeficiency in insulin production. Current treatments for type 1diabetics include insulin injections as well as pancreatic β celltransplantation. Such therapies are risky and often unsuccessful. Forexample, it is difficult to regulate blood glucose with insulinrejections, and thus bouts of hypoglycemia are not uncommon. Insulinalso frequently results in weight gain for the patient. Pancreatic islettransplantation carries all the risks associated with any organtransplantation which include side effects associated with the requiredimmunosuppressive therapies to avoid transplant rejection. Finally, typeI diabetes is most common is children and adolescents with an estimated500,000 to 1 million type I diabetics in the United States alone.

Thus, it is urgent, unmet need for new methods of stimulating pancreaticβ cell regeneration in the treatment of diabetes.

SUMMARY OF THE INVENTION

Provided herein are methods for inducing pancreatic β cell regenerationcomprising administering to a subject in need thereof, a pharmaceuticalcomposition comprising an exendin or an exendin agonist analog in anamount effective to restore normoglycemia, wherein said exendin orexendin agonist analog is administered over a short duration and saideffective amount is from about 3 μg/kg to less than about 100 μg/kg. Inone embodiment, the subject has early onset type I diabetes. In oneembodiment, the subject has type I diabetes. In some embodiments theeffective amount is from about 10 μg/kg to less than about 30 μg/kg.

Further provided herein are methods to achieve a sustained delay in theprogression of, or an amelioration of diabetes in a subject, or a delayin diabetes onset in a subject at risk for diabetes, comprising anabbreviated course of administration of a pharmaceutical compositioncomprising an exendin or an exendin agonist analog in an amounteffective to induce β cell regeneration. In one embodiment, the subjecthas type I diabetes. In some embodiments, the delay in diabetesprogression or the amelioration of diabetes occurs in the absence ofimmunosuppressive therapy. In one embodiment, the effective amount isabout 3 μg/kg to less than about 100 μg/kg twice a day (BID). In someembodiments the effective amount is from about 10 μg/kg to less thanabout 30 μg/kg twice a day. In other embodiments the effective amount isfrom about 3 μg/kg to less than about 30 μg/kg twice a day.

In some embodiments, the exendin is exendin-4. In some embodiment, theexendin agonist analog is a peptide compound of Formula I, Formula II,Formula III, Formula IV, Formula V, Formula VI or Formula VII.

In some embodiments, pancreatic insulin content is restored by at least5%, at least 10%, at least 15%, at least 20%, at least 25%, at least30%, at least 40%, at least 50%, at least 60%, at least 70%, at least80%, at least 90%, at least 95%, or at least 100%.

In some embodiments, the ameriolation of diabetes (e.g., achievingnormoglycemia) after the cessation of exendin or exendin agonist analogtreatment is for at least 1 week, at least 2 weeks, at least 3 weeks, atleast 1 month, at least 2 months, at least 3 months, at least 4 months,at least 5 months, at least 6 months, or at least 1 year.

In some embodiments, the sustained delay in the progression of diabetesis for at least 1 month, at least 2 months, at least 3 months, at least4 months, at least 5 months, at least 6 months, at least 1 year, atleast 18 months, or at least 2 years.

In other embodiments, HbA_(1c) is reduced by at least 0.5%, at least1.0%, at least 1.5%, at least 2.0%, at least 2.5% or at least 3.0%. Infurther embodiments, HbA_(1c) is reduced to less than 7.5%, less than7.0%, less than 6.5%, less than 6.0%, less than 5.5%, less than 5.0%,less than 4.5% or less than 4.0%. In still another embodiment, averagepostprandial blood glucose levels do not exceed 175 mg/dl, 170 mg/dl,165 mg/dl, 160 mg/dl, 155 mg/dl or 150 mg/dl. In another embodiment,overall average daily blood glucose concentration is less than 175mg/dl, less than 165 mg/dl, less than 160 mg/dl, less than 155 mg/dl,less than 150 mg/dl, less than 145 mg/dl, less than 140 mg/dl, less than135 mg/dl, less than 130 mg/dl, less than 125 mg/dl, less than 120mg/dl, less than 110 mg/dl or less than 100 mg/dl.

The pharmaceutical compositions useful in the methods disclosed hereincan be administered by any appropriate means known in the art, forexample, intravenously, transmucosally, intranasally, orally,intramuscularly, subcutaneously, transdermally, by inhalation or bypulmonary administration. In one embodiment, the formulation is asustained release or long acting formulation, that is, the formulationreleases the at least one exendin, exendin agonist, or exendin analogagonist into the body over a given period of time, for example about 1day, about 1 week or about 1 month. In further embodiments, theformulation is administered once a day, every other day, once a week,every other week, every third week, once a month, every other month, orevery third month. In additional embodiments, the formulation furthercomprises a biocompatible polymer and sugar, for example sucrose. In oneparticular embodiment, the formulation is a long-acting formulationcontaining 5% (w/w) of at least one exendin, exendin agonist or exendinanalog agonist, which is administered once a week at a dose equivalentto about from 3 μg/kg to about less than 100 μg/kg BID. In anotherparticular embodiment, the formulation long-acting formulationcontaining 5% (w/w) of at least one exendin, exendin agonist or exendinanalog agonist, is administered once a week at a dose equivalent to fromabout 10 μg/kg to about 30 μg/kg BID. In another particular embodiment,the formulation long-acting formulation containing 5% (w/w) of at leastone exendin, exendin agonist or exendin analog agonist, is administeredonce a week at a dose equivalent to from about 3 μg/kg to about 30 μg/kgBID.

Further provided herein is the use of a pharmaceutical compositioncomprising an amount of at least one exendin, exendin agonist or exendinanalog agonist sufficient to mediate the effects or treat the diseasesor disorders as disclosed herein. Also provided is the use of at leastone exendin, exendin agonist or exendin agonist analog to manufacture amedicament to mediate the effects or treat the diseases or disorders asdisclosed herein.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts the correction of hyperglycemia after diabetes onset inNOD mice using exenatide at various doses.

FIG. 2 depicts the delay in hyperglycemia progression and therestoration of normoglycemia after diabetes onset in NOD mice followinga short course of exenatide.

FIG. 3 shows the reversal of diabetes in NOD mice after exenatideadministration is long lasting.

FIG. 4 illustrates a short course of exenatide results in long-termlower blood glucose in 50% of the exenatide-treated NOD mice.

FIG. 5 depicts a short course of exenatide results long-termnormalization of glycated Hb levels in 50% of the exenatide-treated NODmice.

FIG. 6 shows a short course of exenatide results in long-term increasesin plasma C-peptide levels in 50% of the exenatide-treated NOD mice.

FIG. 7 shows sustained, increased pancreatic insulin levels in 50% ofexenatide-treated NOD mice following a short course of exenatidetreatment.

FIG. 8 shows a partial restoration of pancreatic insulin contentcorrects hyperglycemia in NOD mice.

DESCRIPTION OF THE INVENTION

Pancreatic β cell mass can increase in adult life in response tophysiological stimuli such as pregnancy and obesity as well asregenerate following injury. Suarez-Pinzon et al., Diabetes 54:2596-601(2005). Some studies suggest that β-cell regenerative processes canoccur after diabetes onset in animal models for diabetes. For example,the autoimmune destruction and resulting diabetes in the non-obesediabetic (NOD) mouse is an animal model of human type I diabetes. In NODmice, abrogation of the autoimmune response or transplantation of cellsfrom diabetes-resistant donors results in a restoration of normoglycemiain NOD mice. See, e.g., Maki et al., Proc. Nat'l Acad. Sci. USA89:3434-38 (1992); Kodama et al., Science 302:1223-27 (2003). Providedhere are methods of achieving normoglycemia following diabetes onset bystimulating pancreatic β cell regeneration using an abbreviated courseof exendin or exendin agonist analog, without requiring an abrogation ofthe anti-β cell autoimmune response through immunosuppressive therapies.

Provided herein are methods for inducing pancreatic β cell regenerationcomprising administering to a subject in need thereof, a pharmaceuticalcomposition comprising an exendin or an exendin agonist analog in anamount effective to restore normoglycemia, wherein said exendin orexendin agonist analog is administered over a short duration and saideffective amount is from about 3 μg/kg to less than about 100 μg/kg BID.In one embodiment, the subject has early onset type I diabetes. In oneembodiment, the subject has type I diabetes. In some embodiments theeffective amount is from about 3 μg/kg to about 30 μg/kg BID. In furtherembodiments the effective amount is from about 10 μg/kg to about 30μg/kg BID. In still further embodiments the effective amount is about 10μg/kg BID.

Pancreatic β cell regeneration can be determined by any useful method.For example, β cell regeneration can be determined by analyzingC-peptide levels, Hb1Ac levels, as well as endogenous insulin productionin patients not receiving insulin replacement therapy.

Further provided herein are methods to achieve a sustained delay in theprogression of, or an amelioration of diabetes in a subject, or a delayin diabetes onset in a subject at risk for diabetes, comprising anabbreviated course of administration of a pharmaceutical compositioncomprising an exendin or an exendin agonist analog in an amounteffective to induce β cell regeneration. In one embodiment, the subjecthas type I diabetes. In some embodiments, the delay in diabetesprogression or the amelioration of diabetes occurs in the absence ofimmunosuppressive therapy. In one embodiment, the effective amount isabout 3 μg/kg to less than about 100 μg/kg administered twice a day. Insome embodiments the effective amount is from about 10 μg/kg to about 30μg/kg administered twice a day. In some embodiments the effective amountis from about 3 μg/kg to about 30 μg/kg administered twice a day. Insome embodiments the effective amount is about 10 μg/kg BID.

As used herein, the term “an abbreviated course of administration”includes administration over a limited time period. In some embodiments,the pharmaceutical compositions disclosed herein are administered onceor twice daily for at least one week, at least two weeks, at least threeweeks, at least 4 weeks, at least 5 weeks or at least 6 weeks. In otherembodiments, the pharmaceutical composition disclosed herein isadministered weekly for at least at least one week, at least two weeks,at least three weeks, at least 4 weeks, at least 5 weeks, or at least 6weeks. In still other embodiments, the pharmaceutical compositiondisclosed herein is administered monthly for at least one month, atleast two month, at least three months, at least 4 months, at least 5months, or at least 6 months. It is contemplated that once a patient hasreturned to normoglycemia, the detection of one or more indicator thatdiabetic state may be returning (e.g., detection of hyperglycemia,increased Hb1Ac levels, increased thirst and urination, decreasedC-peptide), subsequent abbreviated administration of the exendin orexendin agonist analog may be employed.

Characteristics of type I diabetes include hyperglycemia, increasedthirst and urine production, increased cholesterol in the blood, andincreased blood triglyceride concentration. In some embodiments, theabsence or improvement of one or more of these characteristics can alsoindicate a delay in diabetes progression or amelioration of diabetes.For example, a decrease in thirst and urine production, a decrease incholesterol in the blood, a reduction in postprandial glycemia, areduction in fasting blood glucose levels, or a decrease in bloodtriglyceride concentration can be indicators of a delay in diabetesprogression or amelioration of diabetes.

In determining the length of the abbreviated course of administration,in some embodiments, the exendin, exendin agonist or exendin analogagonist can be administered according to the methods disclosed hereinfor a period sufficient to achieve a target HbA_(1c), a target fastingglucose level, a target C peptide concentration, a target overall dailyblood glucose concentration, etc. In one embodiment, the exendin,exendin agonist or exendin analog agonist is administered according tomethods disclosed herein for a period sufficient to lower or stabilizefasting glucose levels, reducing or eliminating high or higher thandesired fasting glucose levels.

In some embodiments, the ameriolation of diabetes (e.g., achievingnormoglycemia) after the cessation of exendin or exendin agonist analogtreatment is for at least 1 week, at least 2 weeks, at least 3 weeks, atleast 1 month, at least 2 months, at least 3 months, at least 4 months,at least 5 months, at least 6 months, or at least 1 year.

In some embodiments, the sustained delay in the progression of diabetesis for at least 1 month, at least 2 months, at least 3 months, at least4 months, at least 5 months, at least 6 months, at least 1 year, atleast 18 months, or at least 2 years.

The terms “HbA_(1c)”, “A_(1c)”, “glycated hemoglobin” or“glycohemoglobin” refers to glycosylated hemoglobin.

In other embodiments, HbA_(1c) is reduced by at least 0.5%, at least1.0%, at least 1.5%, at least 2.0%, at least 2.5% or at least 3.0%. Infurther embodiments, HbA_(1c) is reduced to less than 7.5%, less than7.0%, less than 6.5%, less than 6.0%, less than 5.5%, less than 5.0%,less than 4.5% or less than 4.0%. In still another embodiment, averagepostprandial blood glucose levels do not exceed 175 mg/dl, 170 mg/dl,165 mg/dl, 160 mg/dl, 155 mg/dl or 150 mg/dl. In another embodiment,overall average daily blood glucose concentration is less than 175mg/dl, less than 165 mg/dl, less than 160 mg/dl, less than 155 mg/dl,less than 150 mg/dl, less than 145 mg/dl, less than 140 mg/dl, less than135 mg/dl, less than 130 mg/dl, less than 125 mg/dl, less than 120mg/dl, less than 110 mg/dl or less than 100 mg/dl.

The pharmaceutical compositions useful in the methods disclosed hereincan be administered by any appropriate means known in the art, forexample, intravenously, transmucosally, intranasally, orally,intramuscularly, subcutaneously, transdermally, by inhalation or bypulmonary administration. In one embodiment, the formulation is asustained release or long acting formulation, that is, the formulationreleases the at least one exendin, exendin agonist, or exendin analogagonist into the body over a given period of time, for example about 1day, about 1 week or about 1 month. See, e.g., U.S. Provisional Appl.No. 60/709,604, filed 19 Aug. 2005; U.S. Provisional Appl. No.60/779,216, filed 3 Mar. 2006. In further embodiments, the formulationis administered once a day, every other day, once a week, every otherweek, every third week, once a month, every other month, or every thirdmonth. In additional embodiments, the formulation further comprises abiocompatible polymer and sugar, for example sucrose. In one particularembodiment, the formulation is a long-acting formulation containing adrug load of at least one exendin, exendin agonist or exendin analogagonist to achieve a dose equivalent to from about 3 μg/kg to less thanabout 100 μg/kg BID, from about 3 μg/kg to about 30 μg/kg BID, fromabout 10 μg/kg to about 30 μg/kg BID, or about 10 μg/kg BID whenadministered once a week at a dose of 2.0 mg. In another particularembodiment, the formulation long-acting formulation containing drug loadof at least one exendin, exendin agonist or exendin analog agonist toachieve the equivalent of from about 10 μg/kg to about 30 μg/kg BID,from about 3 μg/kg to about 30 μg/kg BID, from about 10 μg/kg to about30 μg/kg BID, or about 10 μg/kg BID when administered once a week at adose of 0.8 mg.

In one embodiment, the exendin, exendin agonist or exendin analogagonist is administered in an extended release, slow release, sustainedrelease or long acting formulation. In one embodiment, the exendin orexendin agonist is administered in a polymer-based sustained releaseformulation. Such polymer-based sustained release formulations aredescribed, for example, in U.S. patent application Ser. No. 11/107,550,filed Apr. 15, 2005.

The exendin, exendin agonist or exendin analog agonist can beadministered by any useful method available. In one embodiment, theexendin or exendin agonist is administered subcutaneously.

In some embodiments, the exendin is exendin-4. In some embodiments, theexendin agonist analog is a peptide compound of Formula I, Formula II,Formula III, Formula IV, Formula V, Formula VI or Formula VII.

The term “exendin” includes naturally occurring exendin peptides thatare found in the salivary secretions of the Gila monster. Exendins ofparticular interest include exendin-3 [SEQ ID NO:2], which is present inthe salivary secretions of Heloderma harridum, and exendin-4 [SEQ IDNO:1], a 39 amino acid petide which is naturally present in the salivarysecretions of Heloderma suspectum (Eng, J., et al., J. Biol. Chem.,265:20259-62, 1990; Eng., J., et al., J. Biol. Chem., 267:7402-05,1992). Animal testing of exendin-4 has shown that its ability to lowerblood glucose persists for several hours. Exendin-4, as it occurs in thesalivary secretions of the Gila monster, is an amidated peptide. Itshould be appreciated, however, that the exendins, exendin agonists andexendin analog agonists for use in the methods described herein are notlimited to the amidated forms, but include that acid form or any otherphysiologically active form of the molecule, including pharmaceuticallyacceptable salts.

Exendin-4 was first thought to be a component of the venom. It nowappears that exendin-4 is devoid of toxicity, and that it instead ismade in salivary glands in the Gila monster. The exendins have somesequence similarity to several members of the glucagon-like peptidefamily, with the highest identity, 53%, being to GLP-1[7-36]NH₂ (Goke,et al., J. Biol. Chem., 268:19650-55, 1993).

Exendin “agonist activity” as used herein means having the biologicalactivity of an exendin, but it is understood that the activity of theactivity can be either less potent or more potent than the nativeexendin. Other exendin agonists include, e.g., chemical compoundsspecifically designed to active that receptor or receptors at which anexendin exerts its effect on pancreatic β cell regeneration.

The term “insulin resistance” as used herein, describes a subnormalbiological response to a given concentration of insulin (i.e., decreasedglucose transport across the cell membrane in response to insulin).

The terms “pharmaceutically” or “pharmacologically acceptable” refer tomolecular entities and compositions that do not produce adverse,allergic, or other untoward reactions when administered to an animal ora human. As used herein, “pharmaceutically acceptable carrier” includesany and all such solvents, dispersion media, coatings, antibacterial andantifungal agents, isotonic and absorption delaying agents and the like.The use of such media and agents for pharmaceutically active substancesis well known in the art. Except insofar as any conventional media oragent is incompatible with the active ingredient, its use in therapeuticcompositions is contemplated. Supplementary active ingredients also canbe incorporated into the compositions.

Certain exendin sequences are compared to the sequence of GLP-1 in Table1.

TABLE 1a. H A E G T F T S D V S S Y L E G Q A A K E F I A W L V K G R (NHb. H S D G T F T S D L S K Q M E E E A V R L F I E W L K N G G P S S G A P P P S (NH₂)c. D L S K Q M E E E A V R L F I E W L K N G G P S S G A P P P S (NH₂)d. H G E G T F T S D L S K Q M E E E A V R L F I E W L K N G G P S S G A P P P S (NH₂)e. H S D A T F T A E Y S K L L A K L A L Q K Y L E S I L G S S T S P R P P S Sf. H S D A T F T A E Y S K L L A K L A L Q K Y L E S I L G S S T S P R P P Sg. H S D A I F T E E Y S K L L A K L A L Q K Y L A S I L G S R T S P P P (NH₂)h. H S D A I F T Q Q Y S K L L A K L A L Q K Y L A S I L G S R T S P P P (NH₂)a = GLP-1(7-36)(NH₂) [SEQ ID NO: 3]. b = exendin 3 (NH₂) [SEQ ID NO: 2].c = exendin 4 (9-39)(NH₂) [SEQ ID NO: 4]. d = exendin 4 (NH₂) [SEQ IDNO: 1]. e = helospectin I [SEQ ID NO: 5]. f = helospectin II [SEQ ID NO:6]. g = helodermin (NH₂) [SEQ ID NO: 7]. h = Q⁸, Q⁹ helodermin (NH₂)[SEQ ID NO: 8].

As used in this specification, by “exendin agonist” is meant a compoundwhich elicits a biological activity of a exendin reference peptide,preferably having a potency better than the exendin reference peptide,or within five orders of magnitude (plus or minus) of potency comparedto the exendin reference peptide, for example, 4, 3, 2, or 1 order ofmagnitude, when evaluated by art-known measures such as receptorbinding/competition studies. In one embodiment, the term refers to acompound which elicits a biological effect similar to that of theexendin reference peptide, for example a compound (1) having activity inglucose lowering and/or pancreatic β regeneration assays similar to theexendin reference peptide, and (2) which optionally binds specificallyin a reference receptor assay or in a competitive binding assay withlabeled exendin reference peptide. Preferably, the agonists will bind insuch assays with an affinity of less than 1 μM, and more preferably withan affinity of less than 1-5 nM. Such agonists may comprise apolypeptide comprising an active fragment of a reference peptide or asmall chemical molecule. In one embodiment, the exendin agonist is apeptide. In another embodiment, exendin agonists do not include GLP-1and variants, analogs and derivatives thereof.

Exendin analog agonists include exendin analogs with agonist activity inwhich one or more naturally occurring amino acids are inserted,eliminated or replaced with another amino acid(s). Exendin analogs arepeptide analogs of exendin-4.

Exendin analogs include peptides that are encoded by polynucleotidesthat express biologically active exendin analogs with agonist activity,as defined herein. Exendin analogs may be peptides containing one ormore amino acid substitutions, additions or deletions, compared withreference exendin, for example, exendin-4. In one embodiment, the numberof substitutions, deletions, or additions is 30 amino acids or less, 25amino acids or less, 20 amino acids or less, 15 amino acids or less, 10amino acids or less, 5 amino acids or less or any integer between 30 and1, inclusive. In one aspect, the substitutions include one or moreconservative substitutions. A “conservative” substitution denotes thereplacement of an amino acid residue by another, biologically active,similar residue. Examples of conservative substitutions include thesubstitution of one hydrophobic residue, such as isoleucine, valine,leucine, or methionine for another, or the substitution of one polarresidue for another, such as the substitution of arginine for lysine,glutamic for aspartic acids, or glutamine for asparagine, and the like.The following table lists illustrative, but non-limiting, conservativeamino acid substitutions.

TABLE 2 ORIGINAL RESIDUE EXEMPLARY SUBSTITUTIONS ALA SER, THR ARG LYSASN HIS, SER ASP GLU, ASN CYS SER GLN ASN, HIS GLU ASP GLY ALA, SER HISASN, GLN ILE LEU, VAL, THR LEU ILE, VAL LYS ARG, GLN, GLU, THR MET LEU,ILE, VAL PHE LEU, TYR SER THR, ALA, ASN THR SER, ALA TRP ARG, SER TYRPHE VAL ILE, LEU, ALA PRO ALA

It is further understood that exendin analogs include the peptidesdescribed herein which have been chemically derivatized or altered, forexample, peptides with non-natural amino acid residues (e.g., taurine,β- and γ-amino acid residues and D-amino acid residues), C-terminalfunctional group modifications, such as amides, esters, and C-terminalketone modifications and N-terminal functional group modifications, suchas acylated amines, Schiff bases, or cyclization, as found, for example,in the amino acid pyroglutamic acid.

Such derivativatized peptides include exendins, exendin agonists andexendin analog agonists conjugated to one or more water soluble polymermolecules, such as polyethylene glycol (“PEG”) or albumin, or gelatin,or fatty acid chains of various lengths (e.g., stearyl, palmitoyl,octanoyl, etc.), or by the addition of polyamino acids, such aspoly-his, poly-arg, poly-lys, and poly-ala. Modifications to theexendins, exendin agonists and exendin analog agonists can also includesmall molecule substituents, such as short alkyls and constrained alkyls(e.g., branched, cyclic, fused, adamantyl), and aromatic groups. Thewater soluble polymer molecules will preferably have a molecular weightranging from about 500 to about 20,000 Daltons.

Such polymer-conjugations and small molecule substituent modificationsmay occur singularly at the N- or C-terminus or at the side chains ofamino acid residues within the sequence of the polypeptides.Alternatively, there may be multiple sites of derivatization along thehybrid polypeptide. Substitution of one or more amino acids with lysine,aspartic acid, glutamic acid, or cysteine may provide additional sitesfor derivatization. See, e.g., U.S. Pat. Nos. 5,824,784 and 5,824,778.In one embodiment, the polypeptides may be conjugated to one, two, orthree polymer molecules.

The water-soluble polymer molecules are typically linked to an amino,carboxyl, or thiol group, and may be linked by N or C terminus, or atthe side chains of lysine, aspartic acid, glutamic acid, or cysteine.Alternatively, the water-soluble polymer molecules may be linked withdiamine and dicarboxylic groups. In one embodiment, the polypeptides areconjugated to one, two, or three PEG molecules through an epsilon aminogroup on a lysine amino acid.

Also included in the present invention are exendin analog sequenceshaving greater than 50% sequence identity, greater than 60% sequenceidentity, greater than 70% sequence identity, greater than 80% sequenceidentity, greater than 90% sequence identity, greater than 95% sequenceidentity, greater than 99% sequence identity or any percent identitybetween 50% and 99%, to a reference exendin peptide, for example, (1)SEQ ID NOS: 1 and 2; and (2) to truncated sequences thereof, whereinsaid truncated sequences contain at least 10 amino acids, at least 20amino acids, at least 25 amino acids, at least 30 amino acids, at least35 amino acids, at least 38 amino acids or N−1 amino acids where Nequals the number of amino acids in the full length or reference peptideor protein. As used herein, sequence identity refers to a comparisonmade between two molecules using standard algorithms well known in theart. The preferred algorithm for calculating sequence identity for thepresent invention is the Smith-Waterman algorithm, where an exendin, forexample SEQ ID NO:1 [i.e., exendin-4], is used as the reference sequenceto define the percentage identity of a comparison peptide over itslength. The choice of parameter values for matches, mismatches, andinsertions or deletions is arbitrary, although some parameter valueshave been found to yield more biologically realistic results thanothers. One preferred set of parameter values for the Smith-Watermanalgorithm is set forth in the “maximum similarity segments” approach,which uses values of 1 for a matched residue and −⅓ for a mismatchedresidue (a residue being either a single nucleotide or single aminoacid). Waterman, Bull. Math. Biol. 46; 473 (1984). Insertions anddeletions (indels), x, are weighted as x_(k)=1+⅓k, where k is the numberof residues in a given insert or deletion. Id.

Novel exendin analogs with agonist activity include those described incommonly owned PCT Application Serial No. PCT/US98/16387 filed Aug. 6,1998, entitled “Novel Exendin Agonist Compounds,” which claims thebenefit of U.S. Patent Application Ser. No. 60/055,404, filed Aug. 8,1997, both of which are herein incorporated by reference.

Other novel exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US98/24210,filed Nov. 13, 1998, entitled “Novel Exendin Agonist Compounds,” whichclaims the benefit of U.S. Provisional Application No. 60/065,442 filedNov. 14, 1997, both of which are herein incorporated by reference.

Still other novel exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US98/24273,filed Nov. 13, 1998, entitled “Novel Exendin Agonist Compounds,” whichclaims the benefit of U.S. Provisional Application No. 60/066,029 filedNov. 14, 1997, both of which are herein incorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US97/14199,filed Aug. 8, 1997, entitled “Methods for Regulating GastrointestinalActivity,” which is a continuation-in-part of U.S. patent applicationSer. No. 08/694,954 filed Aug. 8, 1996, both of which are herebyincorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US98/00449,filed Jan. 7, 1998, entitled “Use of Exendins and Agonists Thereof forthe Reduction of Food Intake,” which claims priority to U.S. ProvisionalApplication No. 60/034,90 filed Jan. 7, 1997, both of which are herebyincorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US01/00719,filed Jan. 9, 2001, entitled “Use of Exendins and Agonists Thereof forModulation of Triglyceride Levels and Treatment of Dyslipidemia,” whichclaims priority to U.S. Provisional Application No. 60/175,365 filedJan. 10, 2000, both of which are hereby incorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US00/00902,filed Jan. 14, 2000, entitled “Novel Exendin Agonist Formulations andMethods of Administration Thereof,” which claims priority to U.S.Provisional Application No. 60/116,380 filed Jan. 14, 1999, both ofwhich are hereby incorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US03/16699,filed May 28, 2003, entitled “Novel Exendin Agonist Formulations andMethods of Administration Thereof,” which claims priority to U.S.application Ser. No. 10/157,224 filed May 28, 2002, both of which arehereby incorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US00/00942,filed Jan. 14, 2000, entitled “Methods of Glucagon Suppression,” whichclaims priority to U.S. Provisional Application No. 60/132,017 filedApr. 30, 1999, both of which are hereby incorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US00/14231,filed May 23, 2000, entitled “Use of Exendins and Agonists Thereof forthe Treatment of Gestational Diabetes Mellitus,” which claims priorityto U.S. application Ser. No. 09/323,867 filed Jun. 1, 1999, both ofwhich are hereby incorporated by reference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US99/02554,filed Feb. 5, 1999, entitled “Inotropic and Diuretic Effects of Exendinand GLP-1,” which claims priority to U.S. Provisional Application No.60/075,122 filed Feb. 13, 1998, both of which are hereby incorporated byreference.

Still other exendin analogs with agonist activity include thosedescribed in commonly owned PCT Application Serial No. PCT/US05/04178filed Feb. 11, 2005, entitled “Hybrid Polypeptides with SelectableProperties”.

Activity as exendin agonists and exendin analogs with agonist activitycan be indicated, for example, by activity in the assays describedherein. Effects of exendins or exendin agonists on pancreatic β cellregeneration can be identified, evaluated, or screened for, using themethods described herein, or other art-known or equivalent methods fordetermining effect on pancreatic β cell regeneration or function.

Certain exemplary exendin analogs with agonist activity include:exendin-4 (1-30) [SEQ ID NO:9: His Gly Glu Gly Thr Phe Thr Ser Asp LeuSer Lys Gln Met Glu Glu Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys AsnGly Gly]; exendin-4 (1-30) amide [SEQ ID NO:10: His Gly Glu Gly Thr PheThr Ser Asp Leu Ser Lys Gln Met Glu Glu Glu Ala Val Arg Leu Phe Ile GluTrp Leu Lys Asn Gly Gly-NH₂]; exendin-4 (1-28) amide [SEQ ID NO:11: HisGly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu Glu Ala ValArg Leu Phe Ile Glu Trp Leu Lys Asn-NH₂]; ¹⁴Leu,²⁵Phe exendin-4 amide[SEQ ID NO:12: His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln LeuGlu Glu Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn Gly Gly Pro SerSer Gly Ala Pro Pro Pro Ser-NH₂]; ¹⁴Leu,²⁵Phe exendin-4 (1-28) amide[SEQ ID NO:13: His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln LeuGlu Glu Glu Ala Val Arg Leu Phe Ile Glu Phe Leu Lys Asn-NH₂]; and¹⁴Leu,²²Ala,²⁵Phe exendin-4 (1-28) amide [SEQ ID NO:14: His Gly Glu GlyThr Phe Thr Ser Asp Leu Ser Lys Gln Leu Glu Glu Glu Ala Val Arg Leu AlaIle Glu Phe Leu Lys Asn-NH₂].

Also included within the scope of the methods provided herein arepharmaceutically acceptable salts of the compounds of formulae I-VIIIand pharmaceutical compositions including said compounds and saltsthereof.

Formula I

Exendin analogs with agonist activity also include those described inU.S. Provisional Application No. 60/065,442, including compounds of theformula (I) [SEQ ID NO:15]:

Xaa₁ Xaa₂ Xaa₃ Gly Xaa₅ Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Xaa₁₁ Xaa₁₂ Xaa₁₃ Xaa₁₄ Xaa₁₅ Xaa₁₆ Xaa₁₇ AlaXaa₁₉ Xaa₂₀ Xaa₂₁ Xaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Xaa₂₆ Xaa₂₇ Xaa₂₈-Z₁;wherein

Xaa₁ is His, Arg or Tyr; Xaa₂ is Ser, Gly, Ala or Thr; Xaa₃ is Ala, Aspor Glu; Xaa₅ is Ala or Thr;

Xaa₆ is Ala, Phe, Tyr or naphthylalanine;

Xaa₇ is Thr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ is Asp or Glu;

Xaa₁₀ is Ala, Leu, Ile, Val, pentylglycine or Met;

Xaa₁₁ is Ala or Ser; Xaa₁₂ is Ala or Lys; Xaa₁₃ is Ala or Gln;

Xaa₁₄ is Ala, Leu, Ile, pentylglycine, Val or Met;

Xaa₁₅ is Ala or Glu; Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala or Glu; Xaa₁₉ isAla or Val; Xaa₂₀ is Ala or Arg; Xaa₂₁ is Ala or Leu;

Xaa₂₂ is Ala, Phe, Tyr or naphthylalanine;Xaa₂₃ is Ile, Val, Leu, pentylglycine, tert-butylglycine or Met;

Xaa₂₄ is Ala, Glu or Asp;

Xaa₂₅ is Ala, Trp, Phe, Tyr or naphthylalanine;

Xaa₂₆ is Ala or Leu; Xaa₂₇ is Ala or Lys; Xaa₂₈ is Ala or Asn; Z₁ is—OH, —NH₂

Gly-Z₂, Gly Gly-Z₂, Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂,Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂ orGly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈- Z₂;

Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently Pro, homoproline, 3Hyp,4Hyp, thioproline, N-alkylglycine, N-alkylpentylglycine orN-alkylalanine; and

Z₂ is —OH or —NH₂;

provided that no more than three of Xaa₃, Xaa₅, Xaa₆, Xaa₈, Xaa₁₀,Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇, Xaa₁₉, Xaa₂₀, Xaa₂₁,Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇ and Xaa₂₈ are Ala.

Exemplary N-alkyl groups for N-alkylglycine, N-alkylpentylglycine andN-alkylalanine include lower alkyl groups preferably of 1 to about 6carbon atoms, more preferably of 1 to 4 carbon atoms.

Exemplary exendin analogs include those wherein Xaa₁ is His or Tyr. Inone embodiment, Xaa₁ is His.

Provided are those compounds wherein Xaa₂ is Gly.

Provided are those compounds wherein Xaa₁₄ is Leu, pentylglycine or Met.

Exemplary compounds are those wherein Xaa₂₅ is Trp or Phe.

Exemplary compounds are those where Xaa₆ is Phe or naphthylalanine;Xaa₂₂ is Phe or naphthylalanine and Xaa₂₃ is Ile or Val.

Provided are compounds wherein Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ areindependently selected from Pro, homoproline, thioproline andN-alkylalanine.

Preferably Z₁ is —NH₂.

Preferably Z₂ is —NH₂.

According to one embodiment, provided are compounds of formula (I)wherein Xaa₁ is His or Tyr, more preferably His; Xaa₂ is Gly; Xaa₆ isPhe or naphthylalanine; Xaa₁₄ is Leu, pentylglycine or Met; Xaa₂₂ is Pheor naphthylalanine; Xaa₂₃ is Ile or Val; Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈are independently selected from Pro, homoproline, thioproline orN-alkylalanine. More preferably Z₁ is —NH₂.

According to one embodiment, exemplary compounds include those offormula (I) wherein: Xaa₁ is His or Arg; Xaa₂ is Gly or Ala; Xaa₃ is Aspor Glu; Xaa₅ is Ala or Thr; Xaa₆ is Ala, Phe or nephthylalaine; Xaa₇ isThr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ is Asp or Glu; Xaa₁₀ is Ala,Leu or pentylglycine; Xaa₁₁ is Ala or Ser; Xaa₁₂ is Ala or Lys; Xaa₁₃ isAla or Gln; Xaa₁₄ is Ala, Leu or pentylglycine; Xaa₁₅ is Ala or Glu;Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala or Glu; Xaa₁₉ is Ala or Val; Xaa₂₀ isAla or Arg; Xaa₂₁ is Ala or Leu; Xaa₂₂ is Phe or naphthylalanine; Xaa₂₃is Ile, Val or tert-butylglycine; Xaa₂₄ is Ala, Glu or Asp; Xaa₂₅ isAla, Trp or Phe; Xaa₂₆ is Ala or Leu; Xaa₂₇ is Ala or Lys; Xaa₂₈ is Alaor Asn; Z₁ is —OH, —NH₂, Gly-Z₂, Gly Gly-Z₂, Gly Gly Xaa₃₁-Z₂, Gly GlyXaa₃₁ Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly AlaXaa₃₆-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂, Gly Gly Xaa₃₁Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈-Z₂; Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈being independently Pro homoproline, thioproline or N-methylalanine; andZ₂ being —OH or —NH₂; provided that no more than three of Xaa₃, Xaa₅,Xaa₆, Xaa₈, Xaa₁₀, Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇,Xaa₁₉, Xaa₂₀, Xaa₂₁, Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇ and Xaa₂₈ are Ala.Especially preferred compounds include those set forth in PCTapplication Serial No. PCT/US98/24210, filed Nov. 13, 1998, entitled“Novel Exendin Agonist Compounds” identified therein as compounds 2-23.

According to another embodiment, provided are compounds where Xaa₁₄ isLeu, Ile, Val or pentylglycine, more preferably Leu or pentylglycine,and Xaa₂₅ is Phe, Tyr or naphthylalanine, more preferably Phe ornaphthylalanine. These compounds will be less susceptive to oxidativedegration, both in vitro and in vivo, as well as during synthesis of thecompound.

Formula II

Exendin analogs with agonist activity also include those described inU.S. Provisional Application No. 60/066,029, including compounds of theformula (II) [SEQ ID NO:16]:

Xaa₁ Xaa₂ Xaa₃ Xaa₄ Xaa₅ Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Xaa₁₁ Xaa₁₂ Xaa₁₃ Xaa₁₄ Xaa₁₅ Xaa₁₆ Xaa₁₇ AlaXaa₁₉ Xaa₂₀ Xaa₂₁ Xaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Xaa₂₆ Xaa₂₇ Xaa₂₈-Z₁;wherein:

Xaa₁ is His, Arg, Tyr, Ala, Norval, Val or Norleu; Xaa₂ is Ser, Gly, Alaor Thr; Xaa₃ is Ala, Asp or Glu; Xaa₄ is Ala, Norval, Val, Norleu orGly; Xaa₅ is Ala or Thr;

Xaa₆ is Ala, Phe, Tyr or naphthylalanine;

Xaa₇ is Thr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ is Ala, Norval, Val,Norleu, Asp or Glu;

Xaa₁₀ is Ala, Leu, Ile, Val, pentylglycine or Met;

Xaa₁₁ is Ala or Ser; Xaa₁₂ is Ala or Lys; Xaa₁₃ is Ala or Gln;

Xaa₁₄ is Ala, Leu, Ile, pentylglycine, Val or Met;

Xaa₁₅ is Ala or Glu; Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala or Glu; Xaa₁₉ isAla or Val; Xaa₂₀ is Ala or Arg; Xaa₂₁ is Ala or Leu;

Xaa₂₂ is Phe, Tyr or naphthylalanine;Xaa₂₃ is Ile, Val, Leu, pentylglycine, tert-butylglycine or Met;

Xaa₂₄ is Ala, Glu or Asp;

Xaa₂₅ is Ala, Trp, Phe, Tyr or naphthylalanine;

Xaa₂₆ is Ala or Leu; Xaa₂₇ is Ala or Lys; Xaa₂₈ is Ala or Asn; Z₁ is—OH, —NH₂

Gly-Z₂, Gly Gly-Z₂, Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂,Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈-Z₂ orGly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Xaa₃₉-Z₂;

Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently Pro, homoproline, 3Hyp,4Hyp, thioproline, N-alkylglycine, N-alkylpentylglycine orN-alkylalanine;

Xaa₃₉ is Ser or Tyr; and

-   -   Z₂ is —OH or —NH₂;    -   provided that no more than three of Xaa₃, Xaa₄, Xaa₅, Xaa₆,        Xaa₈, Xaa₉, Xaa₁₀, Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆,        Xaa₁₇, Xaa₁₉, Xaa₂₀, Xaa₂₁, Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇ and Xaa₂₈        are Ala; and provided also that, if Xaa₁ is His, Arg or Tyr,        then at least one of Xaa₃, Xaa₄ and Xaa₉ is Ala.

Exemplary N-alkyl groups for N-alkylglycine, N-alkylpentylglycine andN-alkylalanine include lower alkyl groups preferably of 1 to about 6carbon atoms, more preferably of 1 to 4 carbon atoms. Suitable compoundsof formula (II) include those described in application Serial No.PCT/US98/24273, filed Nov. 13, 1998, entitled “Novel Exendin AgonistCompounds.”

In one embodiment, such exendin analogs include those wherein Xaa₁ isHis, Ala or Norval. More preferably Xaa₁ is His or Ala. Most preferablyXaa₁ is His.

Provided are those compounds of formula (II) wherein Xaa₂ is Gly.

Provided are those compounds of formula (II) wherein Xaa₃ is Ala.

Provided are those compounds of formula (II) wherein Xaa₄ is Ala.

Provided are those compounds of formula (II) wherein Xaa₉ is Ala.

Provided are those compounds of formula (II) wherein Xaa₁₄ is Leu,pentylglycine or Met.

Exemplary compounds of formula (II) are those wherein Xaa₂₅ is Trp orPhe.

Exemplary compounds of formula (II) are those where Xaa₆ is Ala, Phe ornaphthylalanine; Xaa₂₂ is Phe or naphthylalanine; and Xaa₂₃ is Ile orVal.

Provided are compounds of formula (II) wherein Xaa₃₁, Xaa₃₆, Xaa₃₇ andXaa₃₈ are independently selected from Pro, homoproline, thioproline andN-alkylalanine.

Preferably Z₁ is —NH₂.

Preferably Z₂ is —NH₂.

According to one embodiment, provided are compounds of formula (II)wherein Xaa₁ is Ala, His or Tyr, more preferably Ala or His; Xaa₂ is Alaor Gly; Xaa₆ is Phe or naphthylalanine; Xaa₁₄ is Ala, Leu, pentylglycineor Met; Xaa₂₂ is Phe or naphthylalanine; Xaa₂₃ is Ile or Val; Xaa₃₁,Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected from Pro, homoproline,thioproline or N-alkylalanine; and Xaa₃₉ is Ser or Tyr, more preferablySer. More preferably Z₁ is —NH₂.

According to another embodiment, exemplary compounds include those offormula (II) wherein: Xaa₁ is His or Ala; Xaa₂ is Gly or Ala; Xaa₃ isAla, Asp or Glu; Xaa₄ is Ala or Gly; Xaa₅ is Ala or Thr; Xaa₆ is Phe ornaphthylalanine; Xaa₇ is Thr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ isAla, Asp or Glu; Xaa₁₀ is Ala, Leu or pentylglycine; Xaa₁₁ is Ala orSer; Xaa₁₂ is Ala or Lys; Xaa₁₃ is Ala or Gln; Xaa₁₄ is Ala, Leu, Met orpentylglycine; Xaa₁₅ is Ala or Glu; Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala orGlu; Xaa₁₉ is Ala or Val; Xaa₂₀ is Ala or Arg; Xaa₂₁ is Ala or Leu;Xaa₂₂ is Phe or naphthylalanine; Xaa₂₃ is Ile, Val or tert-butylglycine;Xaa₂₄ is Ala, Glu or Asp; Xaa₂₅ is Ala, Trp or Phe; Xaa₂₆ is Ala or Leu;Xaa₂₇ is Ala or Lys; Xaa₂₈ is Ala or Asn; Z₁ is —OH, —NH₂, Gly-Z₂, GlyGly-Z₂, Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂, Gly Gly Xaa₃₁ SerSer-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly AlaXaa₃₆ Xaa₃₇-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈-Z₂ orGly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Xaa₃₉-Z₂; Xaa₃₁, Xaa₃₆,Xaa₃₇ and Xaa₃₈ being independently Pro homoproline, thioproline orN-methylalanine; and Z₂ being —OH or —NH₂; provided that no more thanthree of Xaa₃, Xaa₅, Xaa₆, Xaa₈, Xaa₁₀, Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄,Xaa₁₅, Xaa₁₆, Xaa₁₇, Xaa₁₉, Xaa₂₀, Xaa₂₁, Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇ andXaa₂₈ are Ala; and provided also that, if Xaa₁ is His, Arg or Tyr, thenat least one of Xaa₃, Xaa₄ and Xaa₉ is Ala. Compounds of formula (II)include those described in application Serial No. PCT/US98/24273, filedNov. 13, 1998, entitled “Novel Exendin Agonist Compounds” as having theamino acid sequence of SEQ ID NOS:5-93 therein.

According to still another embodiment, provided are compounds of formula(II) where Xaa₁₄ is Ala, Leu, Ile, Val or pentylglycine, more preferablyLeu or pentylglycine, and Xaa₂₅ is Ala, Phe, Tyr or naphthylalanine,more preferably Phe or naphthylalanine. These compounds will be lesssusceptible to oxidative degration, both in vitro and in vivo, as wellas during synthesis of the compound.

Formula III

Also useful within the scope of the present invention are narrowergenera of compounds having peptides of various lengths, for examplegenera of compounds which do not include peptides having a length of 28,29 or 30 amino acid residues, respectively. Additionally, the presentinvention includes narrower genera of compounds described in PCTApplication Serial No. PCT/US98/24210, filed Nov. 13, 1998, entitled“Novel Exendin Agonist Compounds” and having particular amino acidsequences, for example, compounds of the formula (III) [SEQ ID NO:17]:

Xaa₁ Xaa₂ Xaa₃ Gly Xaa₅ Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Xaa₁₁ Xaa₁₂ Xaa₁₃ Xaa₁₄ Xaa₁₅ Xaa₁₆ Xaa₁₇ AlaXaa₁₉ Xaa₂₀ Xaa₂₁ Xaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Xaa₂₆ Xaa₂₇ Xaa₂₈-Z₁;wherein:

Xaa₁ is His or Arg; Xaa₂ is Gly or Ala; Xaa₃ is Ala, Asp or Glu; Xaa₅ isAla or Thr;

Xaa₆ is Ala, Phe or naphthylalanine;

Xaa₇ is Thr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ is Asp or Glu;

Xaa₁₀ is Ala, Leu or pentylglycine;

Xaa₁₁ is Ala or Ser; Xaa₁₂ is Ala or Lys; Xaa₁₃ is Ala or Gln;

Xaa₁₄ is Ala, Leu or pentylglycine;

Xaa₁₅ is Ala or Glu; Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala or Glu; Xaa₁₉ isAla or Val; Xaa₂₀ is Ala or Arg; Xaa₂₁ is Ala or Leu;

Xaa₂₂ is Phe or naphthylalanine;Xaa₂₃ is Ile, Val or tert-butylglycine;

Xaa₂₄ is Ala, Glu or Asp; Xaa₂₅ is Ala, Trp, or Phe; Xaa₂₆ is Ala orLeu; Xaa₂₇ is Ala or Lys; Xaa₂₈ is Ala or Asn; Z₁ is —OH, —NH₂,

Gly-Z₂, Gly Gly -Z₂, Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂,Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂ orGly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈- Z₂;

Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected from the groupconsisting of Pro, homoproline, thioproline and N-methylylalanine; and

Z₂ is —OH or —NH₂;

provided that no more than three of Xaa₃, Xaa₅, Xaa₆, Xaa₈, Xaa₁₀,Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇, Xaa₁₉, Xaa₂₀, Xaa₂₁,Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇ and Xaa₂₈ are Ala; and pharmaceuticallyacceptable salts thereof

Formula IV

Additionally, the present invention includes narrower genera of peptidecompounds described in PCT Application Serial No. PCT/US98/24273, filedNov. 13, 1998, entitled “Novel Exendin Agonist Compounds” as havingparticular amino acid sequences, for example, compounds of the formula[IV] [SEQ ID NO:18]:

Xaa₁ Xaa₂ Xaa₃ Xaa₅ Xaa₅ Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Xaa₁₁ Xaa₁₂ Xaa₁₃ Xaa₁₄ Xaa₁₅ Xaa₁₆ Xaa₁₇ AlaXaa₁₉ Xaa₂₀ Xaa₂₁ Xaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Xaa₂₆ Xaa₂₇ Xaa₂₈-Z₁;wherein:

Xaa₁ is His or Ala; Xaa₂ is Gly or Ala; Xaa₃ is Ala, Asp or Glu; Xaa₄ isAla or Gly; Xaa₅ is Ala or Thr;

Xaa₆ is Ala, Phe or naphthylalanine;

Xaa₇ is Thr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ is Ala, Asp or Glu;

Xaa₁₀ is Ala, Leu or pentylglycine;

Xaa₁₁ is Ala or Ser; Xaa₁₂ is Ala or Lys; Xaa₁₃ is Ala or Gln;

Xaa₁₄ is Ala, Leu, Met or pentylglycine;

Xaa₁₅ is Ala or Glu; Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala or Glu; Xaa₁₉ isAla or Val; Xaa₂₀ is Ala or Arg; Xaa₂₁ is Ala or Leu;

Xaa₂₂ is Phe or naphthylalanine;Xaa₂₃ is Ile, Val or tert-butylglycine;

Xaa₂₄ is Ala, Glu or Asp; Xaa₂₅ is Ala, Trp or Phe; Xaa₂₆ is Ala or Leu;Xaa₂₇ is Ala or Lys; Xaa₂₈ is Ala or Asn; Z₁ is —OH, —NH₂,

Gly-Z₂, Gly Gly-Z₂ Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂,Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈-Z₂Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Ser-Z₂;

Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently Pro, homoproline,thioproline, or N-methylylalanine; and

Z₂ is —OH or —NH₂;

provided that no more than three of Xaa₃, Xaa₅, Xaa₆, Xaa₈, Xaa₁₀,Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇, Xaa₁₉, Xaa₂₀, Xaa₂₁,Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇, and Xaa₂₈ are Ala; and provided that, ifXaa₁ is His, Arg or Tyr, then at least one of Xaa₃, Xaa₄ and Xaa₉ isAla; and pharmaceutically acceptable salts thereof.

Exemplary compounds of formula (IV) include those wherein Xaa₁ is His orAla. Preferably, Xaa₁ is His.

Exemplary compounds of formula (IV) include those wherein Xaa₂ is Gly.

Exemplary compounds of formula (IV) include those wherein Xaa₄ is Ala.

Exemplary compounds of formula (IV) include those wherein Xaa₉ is Ala.

Exemplary compounds of formula (IV) include those wherein Xaa₁₄ is Leu,pentylglycine or Met.

Exemplary compounds of formula (IV) include those wherein Xaa₂₅ is Trpor Phe.

Exemplary compounds of formula (IV) include those wherein Xaa₆ is Ala,Phe or naphthylalanine; Xaa₂₂ is Phe or naphthylalanine; and Xaa₂₃ isIle or Val.

Exemplary compounds of formula (IV) include those wherein Z₁ is —NH₂.

Exemplary compounds of formula (IV) include those wherein Xaa₃₁, Xaa₃₆,Xaa₃₇ and Xaa₃₈ are independently selected from the group consisting ofPro, homoproline, thioproline and N-alkylalanine.

Exemplary compounds of formula (IV) include those wherein Z₂ is —NH₂.

Exemplary compounds of formula (IV) include those wherein Z₁ is —NH₂.

Formula V

Also provided are compounds described in PCT application PCT/US98/24210,filed Nov. 13, 1998, entitled “Novel Exendin Agonist Compounds”,including compounds of the formula (V) [SEQ ID NO:19]:

Xaa₁ Xaa₂ Xaa₃ Gly Xaa₅ Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Xaa₁₁ Xaa₁₂ Xaa₁₃ Xaa₁₄ Xaa₁₅ Xaa₁₆ Xaa₁₇ AlaXaa₁₉ Xaa₂₀ Xaa₂₁ Xaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Xaa₂₆/ Xaa₂₇ Xaa₂₈-Z₁;wherein

Xaa₁ is His, Arg or Tyr or 4-imidazopropionyl;

Xaa₂ is Ser, Gly, Ala or Thr;

Xaa₃ is Ala, Asp or Glu;

Xaa₅ is Ala or Thr;

Xaa₆ is Ala, Phe, Tyr or naphthylalanine;

Xaa₇ is Thr or Ser;

Xaa₈ is Ala, Ser or Thr;

Xaa₉ is Asp or Glu;

Xaa₁₀ is Ala, Leu, Ile, Val, pentylglycine or Met;

Xaa₁₁ is Ala or Ser;

Xaa₁₂ is Ala or Lys;

Xaa₁₃ is Ala or Gln;

Xaa₁₄ is Ala, Leu, Ile, pentylglycine, Val or Met;

Xaa₁₅ is Ala or Glu;

Xaa₁₆ is Ala or Glu;

Xaa₁₇ is Ala or Glu;

Xaa₁₉ is Ala or Val;

Xaa₂₀ is Ala or Arg;

Xaa₂₁ is Ala, Leu or Lys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀ straightchain or branched alkanoyl or cycloalkylalkanoyl;

Xaa₂₂ is Phe, Tyr or naphthylalanine;

Xaa₂₃ is Ile, Val, Leu, pentylglycine, tert-butylglycine or Met;

Xaa₂₄ is Ala, Glu or Asp;

Xaa₂₅ is Ala, Trp, Phe, Tyr or naphthylalanine;

Xaa₂₆ is Ala or Leu;

Xaa₂₇ is Lys, Asn, Ala or Lys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀straight chain or branched alkanoyl or cycloalkylalkanoyl;

X₂₈ is Lys, Asn, Ala or Lys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀straight chain or branched alkanoyl or cycloalkylalkanoyl;

Z₁ is —OH, —NH₂,

Gly-Z₂, Gly Gly-Z₂, Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂,Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂ orGly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈- Z₂;

-   -   Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected from        the group consisting of Pro, homoproline, 3Hyp, 4Hyp,        thioproline, N-alkylglycine, N-alkylpentylglycine and        N-alkylalanine; and    -   Z₂ is —OH or —NH₂;        provided that no more than three of Xaa₃, Xaa₅, Xaa₆, Xaa₈,        Xaa₁₀, Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇, Xaa₁₉,        Xaa₂₀, Xaa₂₁, Xaa₂₄, Xaa₂₅, and Xaa₂₆ are Ala. Also within the        scope of the methods provided herein are pharmaceutically        acceptable salts of the compound of formula (V) and        pharmaceutical compositions including said compounds and salts        thereof.

Exemplary exendin analogs of formula (V) include those wherein Xaa₁ isHis, Tyr or 4-imidazopropionyl. More preferably Xaa₁ is His.

Provided are those compounds of formula (V) wherein Xaa₁ is4-imidazopropionyl.

Provided are those compounds of formula (V) wherein Xaa₂ is Gly.

Exemplary compounds of formula (V) are those wherein Xaa₁₄ is Leu,pentylglycine or Met.

Exemplary compounds of formula (V) are those wherein Xaa₂₅ is Trp orPhe.

According to one embodiment, provided are compounds of formula (V)wherein Xaa₆ is Phe or naphthylalanine; and Xaa₂₂ is Phe ornaphthylalanine; and Xaa₂₃ is Ile or Val. More preferably, Z₁ is —NH₂.According to one embodiment, provided are compounds of formula (V)wherein Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected fromthe group consisting of Pro, homoproline, thioproline andN-alkylalanine. Preferably, Z₂ is —NH₂.

Exemplary compounds of formula (V) include those wherein X₂₇ is Lys orLys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀ straight chain or branchedalkanoyl and Xaa₂₈ is Asn or Ala. Preferred compounds of formula (V)include compounds described in PCT application Serial No.PCT/US98/24210, filed Nov. 13, 1998, entitled “Novel Exendin AgonistCompounds” and identified therein as Compound Nos. 62-69.

Provided exendin analogs include those wherein Xaa₁ is His.

Provided are those compounds of formula (V) wherein Xaa₂ is Gly.

Provided are those compounds of formula (V) wherein Xaa₃ is Ala.

Provided are those compounds of formula (V) wherein Xaa₁₄ is Leu,pentylglycine or Met.

Provided compounds of formula (V) are those wherein Xaa₂₅ is Trp or Phe.

Exemplary compounds of formula (V) are those where Xaa₆ is Ala, Phe ornaphthylalanine; Xaa₂₂ is Phe or naphthylalanine; and Xaa₂₃ is Ile orVal.

Provided are compounds of formula (V) wherein Xaa₃₁, Xaa₃₆, Xaa₃₇ andXaa₃₈ are independently selected from Pro, homoproline, thioproline andN-alkylalanine.

Preferably Z₁ is —NH₂.

Preferably Z₂ is —NH₂.

According to one embodiment, provided are compounds of formula (V)wherein Xaa₁ is His or Tyr, more preferably His; Xaa₂ is Ala or Gly;Xaa₆ is Phe or naphthylalanine; Xaa₁₄ is Ala, Leu, pentylglycine or Met;Xaa₂₂ is Phe or naphthylalanine; Xaa₂₃ is Ile or Val; Xaa₃₁, Xaa₃₆,Xaa₃₇ and Xaa₃₈ are independently selected from Pro, homoproline,thioproline or N-alkylalanine; and Xaa₃₉ is Ser or Tyr, more preferablySer. More preferably Z₁ is —NH₂.

According to one embodiment, provided compounds include those of formula(V) wherein: Xaa₁ is His; Xaa₂ is Gly or Ala; Xaa₃ is Ala, Asp or Glu;Xaa₄ is Gly; Xaa₅ is Ala or Thr; Xaa₆ is Phe or naphthylalanine; Xaa₇ isThr or Ser; Xaa₈ is Ala, Ser or Thr; Xaa₉ is Asp or Glu; Xaa₁₀ is Ala,Leu or pentylglycine; Xaa₁₁ is Ala or Ser; Xaa₁₂ is Ala or Lys; Xaa₁₃ isAla or Gln; Xaa₁₄ is Ala, Leu, Met or pentylglycine; Xaa₁₅ is Ala orGlu; Xaa₁₆ is Ala or Glu; Xaa₁₇ is Ala or Glu; Xaa₁₉ is Ala or Val;Xaa₂₀ is Ala or Arg; Xaa₂₁ is Ala or Leu; Xaa₂₂ is Phe ornaphthylalanine; Xaa₂₃ is Ile, Val or tert-butylglycine; Xaa₂₄ is Ala,Glu or Asp; Xaa₂₅ is Ala, Trp or Phe; Xaa₂₆ is Ala or Leu; Xaa₂₇ is Alaor Lys; Xaa₂₈ is Ala or Asn; Z₁ is —OH, —NH₂, Gly-Z₂, Gly Gly-Z₂, GlyGly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly GlyXaa₃₁ Ser Ser Gly-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂, Gly Gly Xaa₃₁Ser Ser Gly Ala Xaa₃₆-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈-Z₂ or Gly Gly Xaa₃₁ SerSer Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Xaa₃₉-Z₂; Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈being independently Pro homoproline, thioproline or N-methylalanine; andZ₂ being —OH or —NH₂; provided that no more than three of Xaa₃, Xaa₅,Xaa₆, Xaa₈, Xaa₁₀, Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇,Xaa₁₉, Xaa₂₀, Xaa₂₁, Xaa₂₄, Xaa₂₅, Xaa₂₆, Xaa₂₇ and Xaa₂₈ are Ala; andprovided also that, if Xaa₁ is His, Arg or Tyr, then at least one ofXaa₃ and Xaa₄ is Ala. Compounds of formula (V) include those describedin PCT application Serial No. PCT/US98/24210, filed Nov. 13, 1998,entitled “Novel Exendin Agonist Compounds” and having the amino acidsequences identified therein as SEQ ID NOS:5-93.

According to one embodiment, provided are compounds of formula (V) whereXaa₁₄ is Ala, Leu, Ile, Val or pentylglycine, more preferably Leu orpentylglycine, and Xaa₂₅ is Ala, Phe, Tyr or naphthylalanine, morepreferably Phe or naphthylalanine. These compounds will be lesssusceptible to oxidative degration, both in vitro and in vivo, as wellas during synthesis of the compound.

Formula VI

Also provided are peptide compounds described in PCT Application SerialNo. PCT/US98/24273, filed Nov. 13, 1998, entitled “Novel Exendin AgonistCompounds”, including compounds of the formula (VI) [SEQ ID NO:20]:

Xaa₁ Xaa₂ Xaa₃ Xaa₄ Xaa₅ Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Xaa₁₁ Xaa₁₂ Xaa₁₃ Xaa₁₄ Xaa₁₅ Xaa₁₆ Xaa₁₇ AlaXaa₁₉ Xaa₂₀ Xaa₂₁ Xaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Xaa₂₆ Xaa₂₇ Xaa₂₈-Z₁;wherein

Xaa₁ is His, Arg, Tyr, Ala, Norval, Val, Norleu or 4-imidazopropionyl;

Xaa₂ is Ser, Gly, Ala or Thr;

Xaa₃ is Ala, Asp or Glu;

Xaa₄ is Ala, Norval, Val, Norleu or Gly;

Xaa₅ is Ala or Thr;

Xaa₆ is Ala, Phe, Tyr or naphthylalanine;

Xaa₇ is Thr or Ser;

Xaa₈ is Ala, Ser or Thr;

Xaa₉ is Ala, Norval, Val, Norleu, Asp or Glu;

Xaa₁₀ is Ala, Leu, Ile, Val, pentylglycine or Met;

Xaa₁₁ is Ala or Ser;

Xaa₁₂ is Ala or Lys;

Xaa₁₃ is Ala or Gln;

Xaa₁₄ is Ala, Leu, Ile, pentylglycine, Val or Met;

Xaa₁₅ is Ala or Glu;

Xaa₁₆ is Ala or Glu;

Xaa₁₇ is Ala or Glu;

Xaa₁₉ is Ala or Val;

Xaa₂₀ is Ala or Arg;

Xaa₂₁ is Ala, Leu or Lys-NH^(ε)—R where R is Lys, Arg, C₁₋₁₀ straightchain or branched alkanoyl or cycloalleyl-alkanoyl;

Xaa₂₂ is Phe, Tyr or naphthylalanine;

Xaa₂₃ is Ile, Val, Leu, pentylglycine, tert-butylglycine or Met;

Xaa₂₄ is Ala, Glu or Asp;

Xaa₂₅ is Ala, Trp, Phe, Tyr or naphthylalanine;

Xaa₂₆ is Ala or Leu;

Xaa₂₇ is Lys, Asn, Lys-NH^(ε)—R or Ala where R is Lys, Arg, C₁-C₁₀straight chain or branched alkanoyl or cycloalkylalkanoyl;

Xaa₂₈ is Lys, Asn, Lys-NH^(ε)—R or Ala where R is Lys, Arg, C₁-C₁₀straight chain or branched alkanoyl or cycloalkylalkanoyl;

Z₁ is —OH, —NH₂,

Gly-Z₂, Gly Gly-Z₂, Gly Gly Xaa₃₁-Z₂, Gly Gly Xaa₃₁ Ser-Z₂,Gly Gly Xaa₃₁ Ser Ser-Z₂, Gly Gly Xaa₃₁ Ser Ser Gly-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇-Z₂,Gly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈-Z₂ orGly Gly Xaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Xaa₃₉-Z₂;

-   -   Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected from        the group consisting of Pro, homoproline, 3Hyp, 4Hyp,        thioproline, N-alkylglycine, N-alkylpentylglycine and        N-alkylalanine;    -   Xaa₃₉ is Ser or Tyr; and    -   Z₂ is —OH or —NH₂;

provided that no more than three of Xaa₃, Xaa₄, Xaa₅, Xaa₆, Xaa₈, Xaa₉,Xaa₁₀, Xaa₁₁, Xaa₁₂, Xaa₁₃, Xaa₁₄, Xaa₁₅, Xaa₁₆, Xaa₁₇, Xaa₁₉, Xaa₂₀,Xaa₂₁, Xaa₂₄, Xaa₂₅, Xaa₂₆, are Ala; and provided also that, if Xaa₁ isHis, Arg, Tyr, or 4-imidazopropionyl then at least one of Xaa₃, Xaa₄ andXaa₉ is Ala.

Exemplary compounds of formula (VI) include those wherein Xaa₁ is His,Ala, Norval or 4-imidazopropionyl. Preferably, Xaa₁ is His, or4-imidazopropionyl or Ala, more preferably His or 4-imidazopropionyl.

Exemplary compounds of formula (VI) include those wherein Xaa₂ is Gly.

Exemplary compounds of formula (VI) include those wherein Xaa₄ is Ala.

Exemplary compounds of formula (VI) include those wherein Xaa₉ is Ala.

Exemplary compounds of formula (VI) include those wherein Xaa₁₄ is Leu,pentylglycine or Met.

Exemplary compounds of formula (VI) include those wherein Xaa₂₅ is Trpor Phe.

Exemplary compounds of formula (VI) include those wherein Xaa₆ is Ala,Phe or naphthylalanine; Xaa₂₂ is Phe or naphthylalanine; and Xaa₂₃ isIle or Val.

Exemplary compounds of formula (VI) include those wherein Z₁ is —NH₂.

Exemplary compounds of formula (VI) include those wherein Xaa₃₁, Xaa₃₆,Xaa₃₇ and Xaa₃₈ are independently selected from the group consisting ofPro, homoproline, thioproline and N-alkylalanine.

Exemplary compounds of formula (VI) include those wherein Xaa₃₉ is Ser.

Exemplary compounds of formula (VI) include those wherein Z₂ is —NH₂.

Exemplary compounds of formula (VI) include those 42 wherein Z₁ is —NH₂.

Exemplary compounds of formula (VI) include those wherein Xaa₂₁ isLys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀ straight chain or branchedalkanoyl.

Exemplary compounds of formula (VI) include those wherein X₂₇ is Lys orLys-NH^(ε)—R, where R is Lys, Arg, C₁-C₁₀ straight chain or branchedalkanoyl and X₂₈ is Asn or Ala.

Other compounds of formula (VI) include those described in PCTApplication Serial No. PCT/US98/24273, filed Nov. 13, 1998, entitled“Novel Exendin Agonist Compounds” as having an amino acid sequenceselected from those identified therein as SEQ ID NOS:95-110.

Formula VII

Compounds useful in the methods provided herein are exendin analogs withagonist activity described in U.S. patent application Ser. No.09/003,869, filed Jan. 7, 1998, entitled “Use of Exendins And AgonistsThereof For The Reduction of Food Intake”, including compounds of theformula (VII) [SEQ ID NO:21]:

Xaa₁ Xaa₂ Xaa₃ Gly Thr Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Ser Lys Gln Xaa₁₄ Glu Glu Glu Ala Val Arg LeuXaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Leu Lys Asn Gly Gly Xaa₃₁Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Xaa₃₉-Zwherein:

Xaa₁ is His, Arg or Tyr;

Xaa₂ is Ser, Gly, Ala or Thr;

Xaa₃ is Asp or Glu;

Xaa₆ is Phe, Tyr or naphthalanine;

Xaa₇ is Thr or Ser;

Xaa₈ is Ser or Thr;

Xaa₉ is Asp or Glu;

Xaa₁₀ is Leu, Ile, Val, pentylglycine or Met;

Xaa₁₄ is Leu, Ile, pentylglycine, Val or Met;

Xaa₂₂ is Phe, Tyr or naphthalanine;

Xaa₂₃ is Ile, Val, Leu, pentylglycine, tert-butylglycine or Met;

Xaa₂₄ is Glu or Asp;

Xaa₂₅ is Trp, Phe, Tyr, or naphthylalanine;

Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently Pro, homoproline, 3Hyp,4Hyp, thioproline, N-alkylglycine, N-alkylpentylglycine orN-alkylalanine;

Xaa₃₉ is Ser, Thr or Tyr; and Z is —OH or —NH₂;

with the proviso that the compound does not have the formula of eitherSEQ ID NOS:1 or 2. Preferred N-alkyl groups for N-alkylglycine,N-alkylpentylglycine and N-alkylalanine include lower alkyl groupspreferably of 1 to about 6 carbon atoms, more preferably of 1 to 4carbon atoms. Also useful in the present invention are pharmaceuticallyacceptable salts of the compounds of formula (VII).

Exemplary exendin analogs include those wherein Xaa₁ is His or Tyr. Morepreferably Xaa₁ is His.

Provided are those compounds wherein Xaa₂ is Gly.

Provided are those compounds wherein Xaa₁₄ is Leu, pentylglycine or Met.

Exemplary compounds include those wherein Xaa₂₅ is Trp or Phe.

Also provided are compounds where Xaa₆ is Phe or naphthalanine; Xaa₂₃ isIle or Val and Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selectedfrom Pro, homoproline, thioproline or N-alkylalanine. PreferablyN-alkylalanine has a N-alkyl group of 1 to about 6 carbon atoms.

According to one embodiment, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are the same aminoacid reside.

Provided are compounds wherein Xaa₃₉ is Ser or Tyr, more preferably Ser.

Preferably Z is —NH₂.

According to one embodiment, provided are compounds of formula (VII)wherein Xaa₁ is His or Tyr, preferably His; Xaa₂ is Gly; Xaa₆ is Phe ornaphthalanine; Xaa₁₄ is Leu, pentylglycine or Met; Xaa₂₂ is Phe ornaphthalanine; Xaa₂₃ is Ile or Val; Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ areindependently selected from Pro, homoproline, thioproline orN-alkylalanine; and Xaa₃₉ is Ser or Tyr, more preferably Ser. Morepreferably Z is —NH₂.

According to another embodiment, exemplary compounds include those offormula (VII) wherein: Xaa₁ is His or Arg; Xaa₂ is Gly; Xaa₃ is Asp orGlu; Xaa₆ is Phe or napthylalanine; Xaa₇ is Thr or Ser; Xaa₈ is Ser orThr; Xaa₉ is Asp or Glu; Xaa₁₀ is Leu or pentylglycine; Xaa₁₄ is Leu orpentylglycine; Xaa₂₂ is Phe or naphthylalanine; Xaa₂₃ is Ile, Val ort-butyltylglycine; Xaa₂₄ is Glu or Asp; Xaa₂₅ is Trp or Phe; Xaa₃₁,Xaa₃₆, Xaa₃₇, and Xaa₃₈ are independently Pro, homoproline, thioproline,or N-methylalanine; Xaa₃₉ is Ser or Tyr: and Z is —OH or —NH₂; with theproviso that the compound does not have the formula of either SEQ IDNOS:1 or 2. More preferably Z is —NH₂.

According to another embodiment, provided are compounds where Xaa₁₄ isLeu, Ile, Val or pentylglycine, more preferably Leu or pentylglycine,and Xaa₂₅ is Phe, Tyr or naphthylalanine, more preferably Phe ornaphthylalanine. These compounds are believed to exhibit advantageousduration of action and to be less subject to oxidative degration, bothin vitro and in vivo, as well as during synthesis of the compound.

Formula VIII

Also provided are compounds described in PCT Application Serial No.PCT/US98/16387, filed Aug. 6, 1998, entitled “Novel Exendin AgonistCompounds”, including compounds of the formula (VIII) [SEQ ID NO:22]:

Xaa₁ Xaa₂ Xaa₃ Gly Thr Xaa₆ Xaa₇ Xaa₈ Xaa₉ Xaa₁₀Ser Lys Gln Xaa₁₄ Glu Glu Glu Ala Val Arg LeuXaa₂₂ Xaa₂₃ Xaa₂₄ Xaa₂₅ Leu Xaa₂₇ Xaa₂₈ Gly GlyXaa₃₁ Ser Ser Gly Ala Xaa₃₆ Xaa₃₇ Xaa₃₈ Xaa₃₉-Zwherein:

Xaa₁ is His, Arg, Tyr or 4-imidazopropionyl;

Xaa₂ is Ser, Gly, Ala or Thr;

Xaa₃ is Asp or Glu;

Xaa₆ is Phe, Tyr or naphthylalanine;

Xaa₇ is Thr or Ser;

Xaa₈ is Ser or Thr;

Xaa₉ is Asp or Glu;

Xaa₁₀ is Leu, Ile, Val, pentylglycine or Met;

Xaa₁₄ is Leu, Ile, pentylglycine, Val or Met;

Xaa₂₂ is Phe, Tyr or naphthylalanine;

Xaa₂₃ is Ile, Val, Leu, pentylglycine, tert-butylglycine or Met;

Xaa₂₄ is Glu or Asp;

Xaa₂₅ is Trp, Phe, Tyr, or naphthylalanine;

Xaa₂₇ is Lys, Asn, or Lys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀ straightchain or branched alkanoyl or cycloalkylalkanoyl;

Xaa₂₈ is Lys, Asn, or Lys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀ straightchain or branched alkanoyl or cycloalkylalkanoyl;

Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently Pro, homoproline, 3Hyp,4Hyp, thioproline, N-alkylglycine, N-alkylpentylglycine orN-alkylalanine;

Xaa₃₉ is Ser, Thr or Tyr; and

Z is —OH or —NH₂;

with the proviso that the compound does not have the formula of eitherSEQ ID NOS:7 or 9. Suitable compounds of formula (VIII) includecompounds described in PCT Application Serial No. PCT/US98/16387, filedAug. 6, 1998, entitled “Novel Exendin Agonist Compounds” having theamino acid sequences of SEQ ID NOS:37-40 therein.

Exemplary exendin analogs of formula (VIII) include those wherein Xaa₁is His, Tyr or 4-imidazopropionyl. More preferably, Xaa₁ is His or4-imidazopropionyl.

Provided are those compounds of formula (VIII) wherein Xaa₂ is Gly.

Provided are those compounds of formula (VIII) wherein Xaa₁₄ is Leu,pentylglycine or Met.

Provided are those compounds of formula (VIII) wherein Xaa₂₅ is Trp orPhe.

Provided are those compounds of formula (VIII) wherein

Xaa₂₇ is Lys or Lys-NH^(ε)—R where R is Lys, Arg, C₁-C₁₀ straight chainor branched alkanoyl and Xaa₂₈ is Asn.

Also provided are compounds of formula (VIII) wherein Xaa₆ is Phe ornaphthylalanine; Xaa₂₂ is Phe or naphthylalanine; Xaa₂₃ is Ile or Valand Xaa₃₁, Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected from Pro,homoproline, thioproline or N-alkylalanine. According to one embodiment,Xaa₃₉ is Ser or Tyr. Provide are compounds wherein Xaa₃₉ is Ser.Preferably, Z is —NH₂.

According to one embodiment, provided are compounds of formula (VIII)wherein Xaa₆ is Phe or naphthylalanine; Xaa₂₂ is Phe or naphthylalanine;Xaa₂₃ is Ile or Val; Xaa₂₇ is Lys or Lys-NH^(ε)—R where R is Lys, Arg,C₁-C₁₀ straight chain or branched alkanoyl, and Xaa₂₈ is Asn; and Xaa₃₁,Xaa₃₆, Xaa₃₇ and Xaa₃₈ are independently selected from Pro, homoproline,thioproline or N-alkylalanine.

In another embodiment, exendins and exendin analogs of the invention donot include the peptides of SEQ ID NOS:7-13. In one embodiment, exendinanalogs include the analogs of Formulas (I-VIII), with the proviso thatthe analogs do not include the peptides of SEQ ID NOs:1-2.

Also useful in the methods provided herein are narrower genera ofcompounds of the disclosed formulas having peptides of various lengths,for example genera of compounds that do not include peptides having alength of 28, 29 or 30 amino acid residues, respectively, or are atleast 31, 32, 33, 34, 35, 36, 37 or 38 amino acids in length.

Exendins, exendin analog agonists and exendin agonists that arepeptides, described herein may be prepared through peptide purificationas described in, for example, Eng, et al., J. Biol. Chem. 265:20259-62,1990; and Eng, et al., J. Biol. Chem. 267:7402-05, 1992, which areincorporated by reference herein. Alternatively, exendins, exendinpeptide agonists and exendin analog agonists may be prepared by methodsknown to those skilled in the art, for example, as described in Raufman,et al., J. Biol. Chem. 267:21432-37, 1992), which is incorporated byreference herein, using standard solid-phase peptide synthesistechniques, for example, using an automated or semiautomated peptidesynthesizer. Typically, using such techniques, an α-N-carbamoylprotected amino acid and an amino acid attached to the growing peptidechain on a resin are coupled at room temperature in an inert solventsuch as dimethylformamide, N-methylpyrrolidinone or methylene chloridein the presence of coupling agents such as dicyclohexylcarbodiimide and1-hydroxybenzotriazole in the presence of a base such asdiisopropylethylamine. The α-N-carbamoyl protecting group is removedfrom the resulting peptide-resin using a reagent such as trifluoroaceticacid or piperidine, and the coupling reaction repeated with the nextdesired N-protected amino acid to be added to the peptide chain.Suitable N-protecting groups are well known in the art, with, forexample, t-butyloxycarbonyl (tBoc) and fluorenylmethoxycarbonyl (Fmoc).

The solvents, amino acid derivatives and 4-methylbenzhydryl-amine resinused in the peptide synthesizer may be purchased from Applied BiosystemsInc. (Foster City, Calif.). The following side-chain protected aminoacids may be purchased from Applied Biosystems, Inc.:BSD-112344.1-Arg(Pmc), Boc-Thr(Bz1), Fmoc-Thr(t-Bu), Boc-Ser(Bzl),Fmoc-Ser(t-Bu), Boc-Tyr(BrZ), Fmoc-Tyr(t-Bu), Boc-Lys(Cl-Z),Fmoc-Lys(Boc), Boc-Glu(Bzl), Fmoc-Glu(t-Bu), Fmoc-His(Trt),Fmoc-Asn(Trt), and Fmoc-Gln(Trt). Boc-His(BOM) may be purchased fromApplied Biosystems, Inc. or Bachem Inc. (Torrance, Calif.). Anisole,dimethylsulfide, phenol, ethanedithiol, and thioanisole may be obtainedfrom Aldrich Chemical Company (Milwaukee, Wis.). Air Products andChemicals (Allentown, Pa.) supplies HF. Ethyl ether, acetic acid andmethanol may be purchased from Fisher Scientific (Pittsburgh, Pa.).

Solid phase peptide synthesis may be carried out with an automaticpeptide synthesizer (Model 430A, Applied Biosystems Inc., Foster City,Calif.) using the NMP/HOBt (Option 1) system and tBoc or Fmoc chemistry(see, Applied Biosystems User's Manual for the ABI 430A PeptideSynthesizer, Version 1.3B Jul. 1, 1988, section 6, pp. 49-70, AppliedBiosystems, Inc., Foster City, Calif.) with capping. Boc-peptide-resinsmay be cleaved with HF (−50° C. to 0° C., 1 hour). The peptide may beextracted from the resin with alternating water and acetic acid, and thefiltrates lyophilized. The Fmoc-peptide resins may be cleaved accordingto standard methods (Introduction to Cleavage Techniques, AppliedBiosystems, Inc., 1990, pp. 6-12). Peptides may also be assembled usingan Advanced Chem Tech Synthesizer (Model MPS 350, Louisville, Ky.).

Peptides may be purified by RP-HPLC (preparative and analytical) using aWaters Delta Prep 3000 system. A C4, C8 or C18 preparative column (10μ,2.2×25 cm; Vydac, Hesperia, Calif.) may be used to isolate peptides, andpurity may be determined using a C4, C8 or C18 analytical column (5μ,0.46×25 cm; Vydac). Solvents (A=0.1% TFA/water and B=0.1% TFA/CH₃CN) maybe delivered to the analytical column at a flowrate of 1.0 ml/min and tothe preparative column at 15 ml/min. Amino acid analyses may beperformed on the Waters Pico Tag system and processed using the Maximaprogram. Peptides may be hydrolyzed by vapor-phase acid hydrolysis (115°C., 20-24 h). Hydrolysates may be derivatized and analyzed by standardmethods (Cohen, et al., The Pico Tag Method: A Manual of AdvancedTechniques for Amino Acid Analysis, pp. 11-52, Millipore Corporation,Milford, Mass. (1989)). Fast atom bombardment analysis may be carriedout by M-Scan, Incorporated (West Chester, Pa.). Mass calibration may beperformed using cesium iodide or cesium iodide/glycerol. Plasmadesorption ionization analysis using time of flight detection may becarried out on an Applied Biosystems Bio-Ion 20 mass spectrometer.Electrospray mass spectroscopy may be carried and on a VG-Trio machine.

Exendins, exendin analog agonists and exendin agonists that are peptidesmay also be prepared using recombinant DNA techniques, using methods nowknown in the art. See, e.g., Sambrook et al., Molecular CLONING: ALABORATORY MANUAL, 2d Ed., Cold Spring Harbor (1989). Alternatively,such compounds may be prepared by homogeneous phase peptide synthesismethods. Non-peptide compounds useful in the present invention may beprepared by art-known methods. For example, phosphate-containing aminoacids and peptides containing such amino acids, may be prepared usingmethods known in the art. See, e.g., Bartlett and Landen, Biorg. Chem.14:356-377 (1986).

Exendins, exendin agonists or exendin analog agonists may be formulatedinto pharmaceutical compositions for administration to subjects,including humans. These pharmaceutical compositions preferably includean amount of an exendin, an exendin agonist or exendin analog agonisteffective to reduce body weight in the subject, and a pharmaceuticallyacceptable carrier.

Pharmaceutically acceptable carriers useful in these pharmaceuticalcompositions include, e.g., ion exchangers, alumina, aluminum stearate,lecithin, serum proteins, such as human serum albumin, buffer substancessuch as phosphates, glycine, sorbic acid, potassium sorbate, partialglyceride mixtures of saturated vegetable fatty acids, water, salts orelectrolytes, such as protamine sulfate, disodium hydrogen phosphate,potassium hydrogen phosphate, sodium chloride, zinc salts, colloidalsilica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-basedsubstances, polyethylene glycol, sodium carboxymethylcellulose,polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers,polyethylene glycol and wool fat.

The compositions of the present invention may be administeredparenterally, orally, by inhalation spray, topically, rectally, nasally,buccally, vaginally or via an implanted reservoir. The term “parenteral”as used herein includes subcutaneous, intravenous, intramuscular,intra-articular, intra-synovial, intrasternal, intrathecal,intrahepatic, intralesional and intracranial injection or infusiontechniques. In one embodiment, the compositions are administered by aninfusion pump or subcutaneous injection of a slow release, extendedrelease, sustained release or long acting formulation. In oneembodiment, subcutaneous injections are administered once a day; onceevery two, three, four, five, or six days; once per week; twice permonth; once a month; every other month or every third month.

Any of the exendins, exendin agonists or exendin analog agonists may beadministered in the acid or amide form. Additionally, any of theexendins, exendin agonists or exendin analog agonists may form saltswith various inorganic and organic acids and bases. Such salts include,without limitation, salts prepared with organic and inorganic acids, forexample, HCl, HBr, H₂SO₄, H₃PO₄, trifluoroacetic acid, acetic acid,formic acid, methanesulfonic acid, toluenesulfonic acid, maleic acid,fumaric acid and camphorsulfonic acid. Salts prepared with basesinclude, without limititation, ammonium salts, alkali metal salts, e.g.,sodium and potassium salts, and alkali earth salts, e.g., calcium andmagnesium salts. Acetate, hydrochloride, and trifluoroacetate salts areparticular examples. The salts may be formed by conventional means, asby reacting the free acid or base forms of the product with one or moreequivalents of the appropriate base or acid in a solvent or medium inwhich the salt is insoluble, or in a solvent such as water which is thenremoved in vacuo or by freeze-drying or by exchanging the ions of anexisting salt for another ion on a suitable ion exchange resin.

Sterile injectable forms of the compositions of this invention may beaqueous or oleaginous suspension. These suspensions may be formulatedaccording to techniques known in the art, using suitable dispersing orwetting agents and suspending agents. The sterile injectable preparationmay also be a sterile injectable solution or suspension in a non-toxicparenterally acceptable diluent or solvent, for example as a solution in1,3-butanediol. Among the acceptable vehicles and solvents that may beemployed are water, Ringer's solution and isotonic sodium chloridesolution. In addition, sterile, fixed oils are conventionally employedas a solvent or suspending medium. For this purpose, any bland fixed oilmay be employed including synthetic mono- or di-glycerides. Fatty acids,such as oleic acid and its glyceride derivatives are useful in thepreparation of injectables, as are natural pharmaceutically-acceptableoils, such as olive oil or castor oil, especially in theirpolyoxyethylated versions. These oil solutions or suspensions may alsocontain a long-chain alcohol diluent or dispersant, such ascarboxymethyl cellulose or similar dispersing agents that are commonlyused in the formulation of pharmaceutically acceptable dosage forms,including emulsions and suspensions. Other commonly used surfactants,such as Tweens, Spans and other emulsifying agents or bioavailabilityenhancers which are commonly used in the manufacture of pharmaceuticallyacceptable solid, liquid, or other dosage forms may also be used for thepurposes of formulation.

Parenteral formulations may be a single bolus dose, an infusion or aloading bolus dose followed with a maintenance dose. These compositionsmay be administered according to any dosage schedule described herein.

The pharmaceutical compositions of this invention may be orallyadministered in any orally acceptable dosage form including, capsules,tablets, aqueous suspensions or solutions. In the case of tablets fororal use, carriers commonly used include lactose and corn starch.Lubricating agents, such as magnesium stearate, are also typicallyadded. For oral administration in a capsule form, useful diluentsinclude lactose and dried cornstarch. When aqueous suspensions arerequired for oral use, the active ingredient is combined withemulsifying and suspending agents. If desired, certain sweetening,flavoring or coloring agents may also be added.

Alternatively, the pharmaceutical compositions of this invention may beadministered in the form of suppositories for rectal administration.These can be prepared by mixing the agent with a suitable non-irritatingexcipient which is solid at room temperature but liquid at rectaltemperature and therefore will melt in the rectum to release the drug.Such materials include cocoa butter, beeswax and polyethylene glycols.

The pharmaceutical compositions of this invention may also beadministered topically. Topical application can be effected in a rectalsuppository formulation (see above) or in a suitable enema formulation.Topically transdermal patches may also be used.

For topical applications, the pharmaceutical compositions may beformulated in a suitable ointment containing the active componentsuspended or dissolved in one or more carriers. Carriers for topicaladministration of the compounds of this invention include, mineral oil,liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene,polyoxypropylene compound, emulsifying wax and water. Alternatively, thepharmaceutical compositions can be formulated in a suitable lotion orcream containing the active components suspended or dissolved in one ormore pharmaceutically acceptable carriers. Suitable carriers include,but are not limited to, mineral oil, sorbitan monostearate, polysorbate60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzylalcohol, and water.

For ophthalmic use, the pharmaceutical compositions may be formulated asmicronized suspensions in isotonic, pH-adjusted sterile saline, or,preferably, as solutions in isotonic, pH-adjusted sterile saline, eitherwith or without a preservative such as benzylalkonium chloride.Alternatively, for ophthalmic uses, the pharmaceutical compositions maybe formulated in an ointment such as petrolatum.

The pharmaceutical compositions of this invention may also beadministered by nasal aerosol or inhalation. Such compositions areprepared according to techniques well-known in the art of pharmaceuticalformulation and may be prepared as solutions in saline, employing benzylalcohol or other suitable preservatives, absorption promoters to enhancebioavailability, fluorocarbons, and/or other conventional solubilizingor dispersing agents.

The amount of exendin, exendin agonist or exendin analog agonist thatmay be combined with the carrier materials to produce a single dosageform will vary depending upon the host treated and the particular modeof administration. The compositions can be formulated so that a dosageof between 0.1-1000 pmoles/kg body weight/minute (when administered byinfusion) of exendin, exendin agonists or exendin analog agonist isadministered to a patient receiving these compositions. In someembodiments of the invention, the dosage is 1-10 pmoles/kg bodyweight/minute (when administered by infusion). In one embodiment thedosage is 0.5-2.0 pmoles/kg/min when administered by intravenousinfusion. The composition may be administered as a single dose, multipledoses, or over an established period of time. In one embodiment, a longacting formulation containing about 5% of an exendin, exendin agonist,or exendin analog agonist, for example, exendin-4 is administered at adose to deliver the equivalent of from about 3 μg/kg to less than about100 μg/kg BID, from about 3 μg/kg to less than about 30 μg/kg BID, fromabout 10 μg/kg to less than about 30 μg/kg BID, or about 10 μg/kg BID ofsaid exendin, exendin agonist or exendin analog agonist. In anotherembodiment, the dosage is about 2.0 mg of a long acting formulationcontaining about a drug load of an exendin, exendin agonist, or exendinanalog agonist for example, exendin-4 sufficient to deliver theequivalent of from about 3 μg/kg to less than about 100 μg/kg BID, fromabout 3 μg/kg to less than about 30 μg/kg BID, from about 10 μg/kg toless than about 30 μg/kg BID, or about 30 μg/kg BID of said exendin,exendin agonist or exendin analog agonist. In additional embodiments,the dosage is about 1.0, 1.25, 1.5, 1.75, 2.25, 2.5, 2.75, 3.0, 3.25,3.5, 3.75, 4.0, 4.25, 4.5, 4.75, or 5.0 mg of a long acting formulationcontaining a percent (drug load) of an exendin, exendin agonist, orexendin analog agonist for example, exendin-4 sufficient to deliver theequivalent of from about 3 μg/kg to less than about 100 μg/kg BID, fromabout 3 μg/kg to less than about 30 μg/kg BID, from about 10 μg/kg toless than about 30 μg/kg BID, or about 30 μg/kg BID of said exendin,exendin agonist or exendin analog agonist. It will be recognized thatthe exact dosage will vary with the percent exendin and the amount ofcarriers and/or excipients in a particular formulation. It is wellwithin the skill of those of ordinary skill in the art to make suchadjustments in order to obtain the desired plasma concentrations of anexendin, exendin agonist or exendin analog agonist described herein.

In order that the invention described herein may be more fullyunderstood, the following examples are set forth. It should beunderstood that these examples are for illustrative purposes only andare not to be construed as limiting this invention in any manner.

Examples Example 1 Reversal of Diabetes in NOD Mice Using Exenatide

Mice.

NOD female mice were purchased from Taconic (Germantown, N.Y.), andhoused and fed under specific pathogen-free conditions. The mice weremonitored daily for diabetes onset by urine testing using Keto-Diastix(Bayer, Etobicoke, Canada). Diabetes onset was diagnosed by the presenceof glucosuria (>6 mmol/l), ketonuria (>1.5 mmol/l) and a 10- to 12-hfasting blood glucose≧9 mmol/l on two consecutive days, measured on aglucose meter (Glucometer Elite; Bayer). Treatments were started within4 to 7 days after diabetes onset in NOD mice aged 10-14 weeks old.

Analysis of Pancreatic Insulin Content.

For insulin assays, half of each pancreas was weighed, minced with finescissors in a small beaker with 1.0 acidified ethanol (75% ethanol, 1.5%12 mmol/l HCl, and 23.5% H₂O), and incubated for 24 hours at 4° C. toextract insulin from tissue. The ethanolic extracts were diluted ininsulin assay buffer, and insulin was measured using a radioimmunoassaykit for rat and mouse insulin (Linco, St. Charles, Mo.).

Pancreatic Histology.

Half of each pancreas was fixed in 10% buffered formalin and embedded inparaffin. Serial sections 4.5 mm thick were cut. Deparaffinized sectionswere stained for b-cells (insulin-positive) by an immunoperoxidasetechnique. The sections were first incubated with a polyclonal guineapig anti-insulin antibody (Dako, Carpenteria, Calif.), then with abiotinylated goat anti-guinea pig antibody (Vector, Burlingame, Calif.)and a strepavidin peroxidase conjugate and substrate kit (InnoGenexiso-IHC DAB kit, San Ramon, Calif.) that stained insulin-positive cellsa golden brown. Coded slides were examined by light microscopy.

Results.

Exenatide (3-100 μg/kg) was given s.c. twice a day for 3 weeks. Bloodglucose (BG) was 12.0±0.4 mmol/L before treatment and rose to 19.5±1.0mmol/L after 3 weeks in vehicle-treated mice. In contrast, BG decreasedprogressively during the 3 weeks of exenatide treatment. Normoglycemia(BG<8 mmol/L) was achieved and lasted for 13 weeks after stoppingtreatments in 4 of 11 mice (36%) treated with 3 μg/kg exenatide, in 6 of12 mice (50%) treated with 10 μg/kg exenatide, and in 3 of 11 mice (27%)of mice treated with 100 μg/kg exenatide. Plasma C-peptide level wasrestored, and pancreative insulin content was partially restored in theNOD ice that remained normoglycemic for 13 weeks (6/12) after only 3weeks of treatment with exenatide at 10 μg/kg. See Table 2 below. Also,pancreatic histology revealed more insulin-stained cells after exenatidetreatment. In summary, a short course of exenatide treatment increasedpancreatic insulin content and reverse hyperglycemia long-term in 50% ofNOD mice with recent-onset diabetes without the need for concomitantimmunotherapy.

TABLE 3 Glucose C-peptide Insulin Mouse Treatment N = mmol/L HbA1C %pmol/ml μg/pancreas NOD Before 5 12.0 ± 0.4 5.5 ± 0.1 1.52 ± 0.36 1.04 ±0.25 NOD Vehicle 9 28.8 ± 0.2 12.5 ± 0.5  0.02 ± 0.01 0.06 ± 0.02 NODExenatide 6  6.3 ± 0.4 3.4 ± 0.2 0.52 ± 0.05 5.66 ± 1.13 NOD-scid None 8 5.4 ± 0.2 3.2 ± 0.1 0.56 ± 0.05 11.81 ± 1.37 

All publications and patent applications cited in this specification areherein incorporated by reference as if each individual publication orpatent application were specifically and individually indicated to beincorporated by reference. Although the foregoing invention has beendescribed in some detail by way of illustration and example for purposesof clarity of understanding, it will be readily apparent to those ofordinary skill in the art in light of the teachings of this inventionthat certain changes and modifications may be made thereto withoutdeparting from the spirit or scope of the appended claims.

We claim:
 1. A method to achieve sustained normoglycemia in a human withdiabetes, comprising administering to a human a pharmaceuticalcomposition comprising an exendin or exendin agonist analog for a periodof time and in an amount effective to achieve normoglycemia in saidhuman, and then ceasing said administering for at least one week,whereby normoglycemia is sustained for said at least one week, whereinthe exendin or exendin agonist analog comprises a sequence havinggreater than 90% sequence identity to SEQ ID NO:1, 2, 3, or
 12. 2. Themethod of claim 1, wherein the sequence has greater than 95% sequenceidentity to SEQ ID NO:1, 2, 3, or
 12. 3. The method of claim 1, whereinthe exendin or exendin agonist analog is conjugated to polyethyleneglycol, albumin, gelatin, or a fatty acid chain.
 4. The method of claim1, wherein the exendin or exendin agonist analog is conjugated to apolyamino acid.
 5. The method of claim 4, wherein the polyamino acid ispoly-His, poly-Arg, poly-Lys, or poly-Ala.